Arterial diameter, a principal modulator of systemic blood pressure and organ blood flow is regulated by changes in the contractility of arterial myocytes. The contractile status of arterial myocytes is determined by several local and global intracellular calcium([Ca2+]i) signals. Ca2+ signal and diameter regulation byinositol 1,4,5-trisphophate (IPS), a phospholipase C-generated second messenger is poorly understood. The conventional view has been that IPS constricts arteries by stimulatingsarcoplasmic reticulum(SR) Ca2+ release in myocytes. Our recently published data indicateda novel mechanism of IPS-induced vasoconstriction that occurred independentlyof SRCa2+ release, and via IPS receptor (IP3R)-and canonicaltransient receptor potential (TRPC) 3-dependent cation current (ICat) activation.However, mechanisms by which IP3R activation stimulates TRPC channelsin arterial myocytes to regulate arterial diameter are unclear. Preliminary data suggest that physical couplingbetween myocyte IPSRs and TRPC3 channels regulates IPS-induced vasoconstriction. Data also indicate that arterial myocyte caveolae facilitate this IP3R-TRPCchannel vasoregulatory mechanism. The central hypothesis ofthis proposal is that in cerebral artery myocytes, caveolae facilitate TRPC3 channel couplingwith IPSRs to mediate vasoconstrictor and IPS-induced membrane depolarization, voltage-dependent Ca2+ channel activation, [Ca2+]i elevation, and contraction. This proposal will investigate 3 specific aims: Aim 1 will test the hypothesis that in cerebral artery myocytes, IP3R to TRPC3 channel physical couplingis required for IPS-inducedICat activation.Aim 2 will test the hypothesis that IP3R to TRPC3 couplingmediates IPS-induced membrane depolarization, [Ca2+]i elevation, and constriction in cerebral arteries. Aim 3 will examine the hypothesis that arterial myocyte caveolae mediate physical and functional coupling of TRPC3 channelsto IPSRs. Experiments to study these aims will integrate techniques performed at molecular, cellular,and intact tissue levels, including Ca2+ imaging,FRET, electrophysiology, pressurized artery myography, and gene suppression. RELEVANCE (See instructions): Mechanisms that regulate blood pressure and flow are incompletely understood. Alterations in arterial contractility are associated with vascular diseases, including stroke and hypertension. This proposal will enhance our knowledge of a novel signaling event that modulates arterial contractility, and will ultimately lead to better insights into alterations that occur in vascular disease. (End of Abstract)